Predator-prey interactions are the interspecific relationships of greatest interest in ecology. Spiders are among the most diverse and ubiquitous terrestrial predators on the planet. Their large dietary breadth is often linked with the development of specific predatory behaviors and morphological adaptations. However, studies on the predatory behavior of spiders have mostly focused on specialist species, leaving behind the ethological variability occurring in generalist species that allow them to respond to the different prey types. For three species of generalist wandering spiders, we searched images of predation events on the Internet to determine the most common prey. Subsequently, the focal predator species were then used in behavioral experiments. Using high-speed videos, handling patterns for different prey types (spider and cricket) were analyzed. Our results show a notable difference in handling patterns between prey types. We found that the spider prey was often rotated around the axis allowing the predator to bite in the ventral region of the prey and thus avoid a counterattack. Contrary, crickets were arbitrarily rotated. Our work may be an indication that these three species of generalist spiders have a preference for manipulating prey differently with a preference to rotate spiders, allowing them to exploit prey with various defensive mechanisms.

Spider venoms have evolved over thousands of years, optimizing feeding and defense mechanisms. Venom components show pharmacological and biotechnological potential, rising interest in their study. However, the isolation of spider toxins for experimental evaluation poses significant challenges. To address this, transcriptomic analysis combined with computational tools has emerged as an appealing approach to characterizing spider venoms. However, many sequences remain unidentified after automatic annotation. In this study, we manually curated a subset of previously unannotated sequences from the Phoneutria nigriventer transcriptome and identified new putative venom components. Our manual analysis revealed 29 % of the analyzed sequences were potential venom components, 29 % hypothetical/uncharacterized proteins, and 17 % cellular function proteins. Only 25 % of the originally unannotated dataset remained without any identification. Most reclassified components were cysteine-rich peptides, including 23 novel putative toxins. We also found glycine-rich peptides (GRP), corroborating the previous description of GRPs in Phoneutria pertyi venom glands. Furthermore, to emphasize the recurrence of the lack of annotation in spider venom glands transcripts, we provide a survey of the percentage of unidentified sequences in several published spider venom transcriptomics studies. In conclusion, our study highlights the importance of manual curation in uncovering novel venom components and underscores the need for improved annotation strategies to fully exploit the medical and biotechnological potential of spider venoms.

Spider venoms are composed, among other substances, of peptide toxins whose selectivity for certain physiological targets has made them powerful tools for applications such as bioinsecticides, analgesics, antiarrhythmics, antibacterials, antifungals and antimalarials, among others. Bioinsecticides are an environmentally friendly alternative to conventional agrochemicals. In this paper, the primary structure of an insecticidal peptide was obtained from the venom gland transcriptome of the ctenid spider Phoneutria depilata (Transcript ID PhdNtxNav24). The peptide contains 53 amino acids, including 10 Cys residues that form 5 disulfide bonds. Using the amino acid sequence of such peptide, a synthetic gene was constructed de novo by overlapping PCRs and cloned into an expression vector. A recombinant peptide, named delta-ctenitoxin (rCtx-4), was obtained. It was expressed, folded, purified and validated using mass spectrometry (7994.61 Da). The insecticidal activity of rCtx-4 was demonstrated through intrathoracic injection in crickets (LD50 1.2 μg/g insect) and it was not toxic to mice. rCtx-4 is a potential bioinsecticide that could have a broad spectrum of applications in agriculture.

Introduction: Spider venoms are a unique source of bioactive peptides, many of which display remarkable biological stability and neuroactivity. Phoneutria nigriventer, often referred to as the Brazilian wandering spider, banana spider or \"armed\" spider, is endemic to South America and amongst the most dangerous venomous spiders in the world. There are 4,000 envenomation accidents with P. nigriventer each year in Brazil, which can lead to symptoms including priapism, hypertension, blurred vision, sweating, and vomiting. In addition to its clinical relevance, P. nigriventer venom contains peptides that provide therapeutic effects in a range of disease models. Methods: In this study, we explored the neuroactivity and molecular diversity of P. nigriventer venom using fractionation-guided high-throughput cellular assays coupled to proteomics and multi-pharmacology activity to broaden the knowledge about this venom and its therapeutic potential and provide a proof-of-concept for an investigative pipeline to study spider-venom derived neuroactive peptides. We coupled proteomics with ion channel assays using a neuroblastoma cell line to identify venom compounds that modulate the activity of voltage-gated sodium and calcium channels, as well as the nicotinic acetylcholine receptor. Results: Our data revealed that P. nigriventer venom is highly complex compared to other neurotoxin-rich venoms and contains potent modulators of voltage-gated ion channels which were classified into four families of neuroactive peptides based on their activity and structures. In addition to the reported P. nigriventer neuroactive peptides, we identified at least 27 novel cysteine-rich venom peptides for which their activity and molecular target remains to be determined. Discussion: Our findings provide a platform for studying the bioactivity of known and novel neuroactive components in the venom of P. nigriventer and other spiders and suggest that our discovery pipeline can be used to identify ion channel-targeting venom peptides with potential as pharmacological tools and to drug leads.

The transcriptome of the venom glands of the Phoneutria depilata spider was analyzed using RNA-seq with an Illumina protocol, which yielded 86,424 assembled transcripts. A total of 682 transcripts were identified as potentially coding for venom components. Most of the transcripts found were neurotoxins (156) that commonly act on sodium and calcium channels. Nevertheless, transcripts coding for some enzymes (239), growth factors (48), clotting factors (6), and a diuretic hormone (1) were found, which have not been described in this spider genus. Furthermore, an enzymatic characterization of the venom of P. depilata was performed, and the proteomic analysis showed a correlation between active protein bands and protein sequences found in the transcriptome. The transcriptomic analysis of P. depilata venom glands show a deeper description of its protein components, allowing the identification of novel molecules that could lead to the treatment of human diseases, or could be models for developing bioinsecticides.

UNASSIGNED: Spider venoms induce different physio-pharmacological effects by binding with high affinity on molecular targets, therefore being of biotechnological interest. Some of these toxins, acting on different types of ion channels, have been identified in the venom of spiders of the genus Phoneutria, mainly from P. nigriventer. In spite of the pharmaceutical potential demonstrated by P. nigriventer toxins, there is limited information on molecules from venoms of the same genus, as their toxins remain poorly characterized. Understanding this diversity and clarifying the differences in the mechanisms of action of spider toxins is of great importance for establishing their true biotechnological potential. This prompted us to compare three different venoms of the Phoneutria genus: P. nigriventer (Pn-V), P. eickstedtae (Pe-V) and P. pertyi (Pp-V).
UNASSIGNED: Biochemical and functional comparison of the venoms were carried out by SDS-PAGE, HPLC, mass spectrometry, enzymatic activities and electrophysiological assays (whole-cell patch clamp).
UNASSIGNED: The employed approach revealed that all three venoms had an overall similarity in their components, with only minor differences. The presence of a high number of similar proteins was evident, particularly toxins in the mass range of ~6.0 kDa. Hyaluronidase and proteolytic activities were detected in all venoms, in addition to isoforms of the toxins Tx1 and Tx2-6. All Tx1 isoforms blocked Nav1.6 ion currents, with slight differences.
UNASSIGNED: Our findings showed that Pn-V, Pe-V and Pp-V are highly similar concerning protein composition and enzymatic activities, containing isoforms of the same toxins sharing high sequence homology, with minor modifications. However, these structural and functional variations are very important for venom diversity. In addition, our findings will contribute to the comprehension of the molecular diversity of the venoms of the other species from Phoneutria genus, exposing their biotechnological potential as a source for searching for new active molecules.

Molecules that selectively act on N-methyl-D-aspartate (NMDA) receptors may have a multidirectional effect by modulating the activity of NMDARs, affecting their active sites as well as by changing the composition of their subunits. The results of the clinical trials conducted so far in mood disorders and schizophrenia indicate that such agents may become new effective drugs for the treatment of these diseases. Number of spider neurotoxins e.g. ctenitoxins extracted from Phoneutria sp. venom act as potent and selective NMDAR blockers that do not disturb cortical and hippocampal glutamate signaling, LTP generation and synaptic neurochemistry. Possibly this intriguing kind of promising neuroregulatory peptides and polyamines can be clinically applicable in a wide spectrum of neuropsychiatric disorders, including epilepsy, neurotrauma and ischemic injuries. These novel medications can potentially be helpful in the future treatment of stroke and several neurodegenerative diseases.

Arachnids are the most abundant land predators. Despite the importance of their functional roles as predators and the necessity to understand their diet for conservation, the trophic ecology of many arachnid species has not been sufficiently studied. In the case of the wandering spider, Phoneutria boliviensis F. O. Pickard-Cambridge, 1897, only field and laboratory observational studies on their diet exist. By using a DNA metabarcoding approach, we compared the prey found in the gut content of males and females from three distant Colombian populations of P. boliviensis. By DNA metabarcoding of the cytochrome c oxidase subunit I (COI), we detected and identified 234 prey items (individual captured by the spider) belonging to 96 operational taxonomic units (OTUs), as prey for this wandering predator. Our results broaden the known diet of P. boliviensis with at least 75 prey taxa not previously registered in fieldwork or laboratory experimental trials. These results suggest that P. boliviensis feeds predominantly on invertebrates (Diptera, Lepidoptera, Coleoptera, and Orthoptera) and opportunistically on small squamates. Intersex and interpopulation differences were also observed. Assuming that prey preference does not vary between populations, these differences are likely associated with a higher local prey availability. Finally, we suggest that DNA metabarcoding can be used for evaluating subtle differences in the diet of distinct populations of P. boliviensis, particularly when predation records in the field cannot be established or quantified using direct observation.

Phoneutria nigriventer venom (PNV) is a complex mixture of toxins exerting multiple pharmacological effects that ultimately result in severe local pain at the site of the bite. It has been proposed that the PNV-induced pain is mediated by both peripheral and central mechanisms. The nociception triggered by PNV is peripherally mediated by the activation of B2, 5-HT4, NMDA, AMPA, NK1, and NK2 receptors, as well as TTXS-Na+, ASIC, and TRPV1 channels. The activation of tachykinin, glutamate and CGRP receptors along with the production of inflammatory mediators are, at least partially, responsible for the central component of pain. Despite its well established pro-nociceptive properties, PNV contains some toxins with antinociceptive activity, which have been studied in the last few years. The toxins ω-CNTX-Pn4a, ω-CNTX-Pn2a, ω-CNTX-Pn3a, κ-CNTX-Pn1a, U7-CNTX-Pn1a, δ-CNTX-Pn1a, and Γ-CNTX-Pn1a from PNV, as well as the semi-synthetic peptide PnPP-19 have been tested in different experimental models of pain showing consistent antinociceptive properties. This review aims to discuss the pro- and antinociceptive actions of PNV and its toxins, highlighting possible mechanisms involved in these apparently dualistic properties.

BACKGROUND: Phoneutria nigriventer spider venom contains several cysteine-rich peptide toxins that act on different ion channels. Despite extensive studies on its venom and description of cDNA sequences of several of its toxin precursors, the gene structure of these toxins remains unknown.
METHODS: Genomic regions encoding the precursors of three previously characterized P. nigriventer toxins - PnTx1, PnTx2-5 and PnTx4(5-5) - were amplified by PCR using specific primers. PCR fragments were cloned and sequenced. Obtained sequences were compared with their corresponding cDNA sequences.
RESULTS: The size of PCR fragments obtained and sequences corresponding to genomic regions encoding for the toxin precursors matched their cDNA sequences.
CONCLUSIONS: Despite a few nucleotide substitutions in the genomic regions encoding for the toxin precursors when compared with cDNA sequences, the results of the present work indicate that P. nigriventer toxins do not contain introns in their genes sequences.